Pancreatic ductal adenocarcinoma (PDAC) is a devastating malignant disease with a median survival of less than 6 months and an overall 5-year survival rate of 1-4% (Pierantoni, Pagliacci et al. 2008). Lack of early diagnosis and effective systemic treatment are major reasons that account for these dismal survival rates. Morphologic and genetic analyses have implicated pancreatic intraepithelial neoplasm (PanIN) as a precursor lesion of human PDAC. PanINs appear to evolve in a stepwise manner through stages (PanIN1A, 1B, 2, 3) that display increasing cellular atypia and accumulate clonal mutations or aberrant expression of oncogenes or tumor suppressor genes such as K-Ras, p16, p53, and DPC4/SMAD4 in the course of progression to PDAC (Goggins, Kern et al. 1999). However, drugs that target these molecular abnormalities have not yet translated into improved clinical responses (Strimpakos, Saif et al. 2008). The aggressive nature of PDAC is featured by its early invasion and metastasis. This process in pancreatic cancer is also characterized by the epithelial-mesenchymal transition (Natalwala, Spychal et al. 2008). However, little is known about the molecular mechanisms underlying the early invasion and metastasis of PDAC. A better understanding of such mechanisms is believed to be essential for the identification of new targets for more effective treatments of PDAC. Thus, the identification of additional biologic pathways that contribute to the pathogenesis of PDAC will enhance our understanding of its biology and will facilitate the development of improved therapeutic interventions for the treatment of this deadly disease.
Cancer immunotherapy is an emerging approach for the treatment of pancreatic cancer. Early clinical studies are also providing critical human reagents for developing methods to identify new candidate proteins and biologic pathways. In particular, immunized lymphocytes and sera are being used to develop functional genomic and proteomic approaches for identifying those proteins that are relevant to the cancer. We have developed an allogeneic, GM-CSF secreting pancreatic cancer vaccine approach (Jaffee, Hruban et al. 2001). Phase I and II trials evaluating this vaccine in patients with resected PDAC have demonstrated both clinical and immunologic responses (Jaffee, Hruban et al. 2001; Laheru, Lutz et al. 2008) (Lutz et al. Manuscript submitted). This whole-cell vaccine based immunotherapy approach has provided critical lymphocyte reagents to develop a functional genomic approach for identifying novel pancreatic cancer antigens that are the targets of the immune response. As a result, we have reported the identification of mesothelin as a T cell target expressed by PDACs of patients treated with the pancreatic cancer vaccine who also developed other evidence of immune responses. (Thomas, Santarsiero et al. 2004). Mesothelin is a GPI linked cell surface glycoprotein expressed by the majority of PDACs and appears to be an adhesion molecule involved in tumor metastasis (Argani, Iacobuzio-Donahue et al. 2001). A number of therapeutic interventions targeting this molecule are currently in phase I testing (Hassan and Ho 2008)
Humoral immune response is an important and integrated part of the immune mechanisms by which a host defends itself against pathogen assault. Antibodies generated from vaccinations are a major factor that has protected generations of children from deadly infectious diseases. On the other hand, antibodies can also mediate pathogenesis in many autoimmune diseases in which the antibodies target cellular components of the host, i.e., auto-antigens, which under normal physiological conditions are tolerated by host immune system (1). As in autoimmune diseases, antigens in cancer come from within. Indeed, a common repertoire of autoantibodies was found to be shared by cancer and autoimmune disease patients (2). In addition, a majority of these autoantibodies are directed against intracellular components, which leads to the assumption that autoantigens in both cancer and autoimmune diseases emerge from damaged cells (3). Yet, autoantibodies can be detected at the very early pre-malignant stage of cancer development when there is no obvious cancer cell death or inflammation (4). It appears that aberrant gene expression, post-translational modification, and/or protein re-localization in cancer cells gives rise to antigens that are expressed either (a) to a greater extent than in normal cells or (b) as “altered” molecules absent in corresponding normal cells, or (c) in cell compartments where they are not supposed to be under normal conditions (ectopic expression), e.g., nuclear or cytoplasmic proteins appearing on the cell surface and non-secreted proteins being secreted to extracellular milieu. These molecules are collectively named tumor associated antigens (TAAs).
Early attempts by Old and co-workers to analyze humoral responses to cancer cell surface antigens, used an autologous typing approach in which cultured tumor cells were tested for reactivity with serum samples from the same patient (5-9). The analysis of over 200 patients with melanoma, leukemia, malignant brain tumor and renal cancer revealed that sera from a majority of the patients reacted to both tumor and normal cells. In a few rare cases, serum reactivity was directed against antigens present only on autologous tumor cells (unique antigens) or shared on some allogeneic tumor cells (shared tumor antigens). The true nature of the unique or specific tumor antigens was not well defined. One shared antigen Old and co-workers discovered, gangliosides, was investigated further. All ganglioside-reactive antibodies detected in cancer patients were of the IgM class; this was also the immunoglobulin class induced by vaccination of melanoma patients with pure or modified gangliosides (10, 11), which means that no memory response was developed. Nonetheless, ganglioside GM2 antibody production in melanoma patients was associated with a prolonged disease-free interval and survival (12). With the advent of new assay methodologies, such as serological identification of antigens by recombinant expression cloning (SEREX), serological proteome analysis (SERPA) and protein microarrays, a large number of antibody-reactive tumor associated antigens (TAAs) were identified. For example, over 2700 sequences were identified by SEREX alone and have been deposited in the Cancer Immunome Database (13). However, the clinical significance of these TAAs and their antibodies largely remains unclear. Spontaneous antibodies against several TAAs such as p53, NY-ESO-1, and survivin were found more frequently in high-grade tumors and appeared to be associated with disease relapse and poor survival (14-17). High titers of Her2/neu and MUC1 antibodies could be detected at early stages and appeared to be markers of a favorable prognosis (18-20). On the other hand, regardless of whether the respective spontaneous antibodies predict poor or favorable prognosis, vaccination with NY-ESO-1 or MUC1 has been shown to readily induce both humoral and T cell responses that are beneficial to patients (21-23).
The majority of the TAAs identified thus far have been in melanoma and a few other cancers. For cancers like pancreatic cancer, in which TAAs are poorly characterized, whole tumor cell vaccines present a good source for investigating TAAs immunotherapy. Vaccination with irradiated whole tumor cells expressing granulocyte-macrophage colony-stimulating factor (GM-CSF) can induce potent systemic immune responses that are capable of eradicating tumors (24). GM-CSF-secreting tumor vaccines can induce both CD4+ and CD8+ T cell-mediated antitumor responses and a broad range of antibody responses (25-27). Recently, we completed a phase II trial on an allogeneic GM-CSF-secreting pancreatic tumor vaccine in 60 patients with stage 1, 2, and 3-resected pancreatic ductal adenocarcinoma (PDA) (28). Twelve of the 60 vaccinated patients have survived greater than 3 years and shown a favorable clinical response.
There is a continuing need in the art to identify targets that are physiologically relevant to the immune system of human cancer patients.